The overall goal of this application is to determine the molecular basis for TCR antagonism and to investigate the in vivo effects and significance of antagonism in normal T-cell physiology. Under Specific Aim 1, molecular mechanisms of TCR antagonism will be explored. This will include an analysis of the role of CD8 in agonist/antagonist discrimination. Preliminary data suggest that lowering CD8 levels converts an agonist into antagonist and that the presence of CD8 increases TCR binding affinity for both agonist and antagonist ligands. In addition, it will be determined whether T-cell tyrosine kinases (Lck, Fyn, ZAP-70) are activated during antagonist interactions, and the effect of over-expression of specific T-cell tyrosine kinases on agonist/antagonist discrimination will be explored. Preliminary data indicate marked tyrosine phosphorylation with agonist but not with antagonist ligands. Finally, the hypothesis that TCR antagonists inhibit T-cell responses by inducing a dominant negative signal will be tested using T-cells that bear two TCRs with different specificities. Preliminary data support the feasibility of the latter approach. Specific Aim 2 focuses on the identification and characterization of natural TCR antagonists. Exciting preliminary data provide evidence for at least one such naturally occurring antagonist peptide isolated from MHC molecules. The origin, tissue distribution, and T-cell specificity of these natural peptides will be studied. In Specific Aim 3, TCR antagonists will be analyzed in vivo. The effect of artificial TCR antagonists on T-cell development and reactivity will be examined using well characterized ligands and established tg mice (OVA-tcr-1 system) and knockout mice (RAG-2, TAP-1). This will include an analysis of thymic selection, perpherial T-cell longevity and migration, and T-cell antigen reactivity.